Week 13: Cell-cycle Checkpoints and Cancer Intro Flashcards
1
Q
Cell cycle control system
A
complex network of regulatory proteins that act as a series of binary (on/off), irreversible, biochemical switches
- Ensures that the cell cycle events occur in a proper sequence and each event is completed before the next begins
2
Q
checkpoints
A
major regulatory transitions which arrest the cell cycle if previous events have not been completed or environment is unfavorable ⇒ series of mechanisms to make sure you don’t go to the next cell cycle step unless you confirm the current or previous events are completely done
3
Q
what are the cell cycle checkpoints?
A
- G1
- S
- G2
- M
4
Q
G1 checkpoint general info
A
growth phase at the start which is a gap phase right after mitosis (M phase)
- You have 2 daughter cells each starting G1
- Cells will probably be small ⇒ want to make sure the cell is big enough to enter the next cycle
- Also makes sure the cell has enough nutrition and is in a good environment to divide
5
Q
S general info
A
begin replication
6
Q
G2 checkpoint general info
A
waiting phase before you start dividing
- Make sure that you have doubled the genetic information completely
7
Q
M checkpoint general info
A
undergo division and cytokinesis
8
Q
G1 checkpoint
A
sense the cell size, physiological state of the cell, and environmental conditions
- Nutrition, growth factors, mitogens, etc.
9
Q
how is G1-S transition regulated?
A
mitogens stimulates G1-S transition
- Activation of MAP kinase results in an increased production of gene regulatory proteins such as Myc (downstream)
10
Q
Myc
A
transcription factor that triggers increased G1-Cdk activity via gene activation
- helps promote cell cycle entry and stimulates gene transcription that increases cell growth
11
Q
Delayed genes
A
downstream of Myc there are many target genes
12
Q
what is S-phase initiation controlled by?
A
E2F and Retinoblastoma protein (rb)
13
Q
what is the activation pathway of Myc? (5)
A
- mitogen binds a ligand to a transmembrane protein mitogen receptor
- the protein stimulates Ras
- activation of transcription regulatory proteins occurs in the cytosol
- transcription regulatory proteins enter the nucleus and bind to a gene sequence
- Myc is transcribed and translated
14
Q
what are the 2 molecules for G1/S transition?
A
- E2F
- Rb
15
Q
E2F protein
A
transcriptional factor required for the transcription of S-phase genes (G1/S and S-cyclins, DNa polymerase, E2F
- Once you activate E2F you get more E2F
16
Q
Rb protein
A
regulatory protein that binds and inhibits E2F
17
Q
what is the process of the G1-cdk to the G1/S transition? (5)
A
- Mac delayed response of gene expression activates G1-cdk
- G1-Cdk phosphorylates Rb (phosphorylated Rb is inactive) and E2F is released and activated
- active E2F activates positive feedback and S-phase gene transcription for G1/S-cyclin and S-cyclin
- G1/S transition is activated and cyclins activate G1/S-cdk as well as S-Cdk
- S-Cdk activates DNA synthesis for replication of chromosomes
18
Q
what does E2F promote?
A
more E2F production through positive feedback
19
Q
G1/S-Cdk phosphorylates what?
A
Rb to inactivate it and stop binding with E2F
20
Q
S-cdk phosphorylates what?
A
Rb to inactivate it and stop binding with E2F
21
Q
what are the key molecules for S phase?
A
DNA polymerase for replication
- All other molecules used are downstream of E2F
22
Q
T/F we know a lot about the G2/M checkpoint? What do we know?
A
we do not know much
- Unreplicated DNA inhibits Cdc25 is all that we know but not how this is done
23
Q
G2/M checkpoint
A
ensures that DNA is replicated properly ⇒ initiation of mitosis cannot occur until DNA replication is completed
- entry into mitosis is blocked by incomplete DNA replication
24
Q
Hydroxyurea
A
inhibits DNA synthesis by halting it in S phase
25
caffeine
blocks G2/M checkpoint mechanism
26
what was the experiment done with the G2/M checkpoint?
hydroxyurea or caffeine (chemicals) cause inhibition of G2/M because no active Cdc25
- When you add the additional toxin caffeine at a high dose, it also inhibits the checkpoint which means it wrongly will undergo mitosis and result in broken sets of chromosomes in daughter cells
- They will die if lucky but may become cancerous
27
what does the combination of hydroxyurea and caffeine do?
induces incomplete DNA replication and broken/incomplete sets of chromosomes which causes suicidal mitosis
28
metaphase checkpoint (kinetochore attachment)
ensures that all chromosomes have attached to the spindle
- Sister chromatid separation cannot occur until all chromosomes are properly attached to the spindle and unattached chromosomes block sister chromatid separation
29
what do unattached kinetochores block?
APC/C-cdc20 complex
30
spindle assembly checkpoint
ensures that cells do not enter anaphase until all chromosomes are properly attached to the spindles
- Improperly attached kinetochores send out a diffusible signal that blocks Cdc-20-APC/C activation
31
what do unattached kinetochores catalyze conformational changes in?
Mad2
- Mad2 (with other proteins) binds and inhibits Cdc20-APC/C
32
DNA damage checkpoint
arrests the cell cycle when DNA is damaged to give time for DNA repair mechanisms ⇒ cells sense the damage
33
what are the 2 ways DNA damage checkpoint works?
1. G1 checkpoint prevents entry into S phase
2. G2 checkpoint prevents entry into mitosis
34
DNA Damage checkpoint (G1)
DNA damage triggers the activation of a set of protein kinases ⇒ ATM, ATR, Chk1, Chk2
35
ATM and ATR
activate the actual players against DNA damage called Chk1 and 2
36
Chk1/2
when active these phosphorylate p53 as their major target
- you don’t find p53 in normal cells because it is always synthesized and degraded
37
p53
is usually bound and ubiquitylated by Mdm2 (Ubi ligase) and degraded in proteasomes
- Phosphorylation of p53 blocks Mdm2 binding, resulting in p53 accumulation because it is unubiquitylated ⇒ done by Chk1 and Chk2
- Active p53 enters the nucleus and binds the regulatory region of the p21 gene (CKI protein) and stimulates expression
38
p21 protein (CDK inhibitor protein)
binds to and inactivates G1/S-Cdk and S-Cdk arresting cells in G1
39
DNA damage checkpoint (G2)
when DNA is damaged, Chk1 and Chk2 phosphorylate and inhibit Cdc25 blocking progression into mitosis
- cdc25 activates M-cdk
40
what do growth factor signals activate? (growth factors include mitogen)
mTORC1 kinase
41
aside from growth factors what also stimulates activation of mTORC1?
cytosolic amino acids
42
mitogen
promotes division
43
growth factors
increase cell size
44
mTORC (mechanistic target of rapamycin)
when active it stimulates protein synthesis, lipid synthesis, and reduces protein turnover ⇒ controls cell size
- kinase that integrates information coming from growth factors such as insulin like growth factors conveying information of nutrition
- nutrition like amino acids, etc. get integrated
45
what does mTORC1 do downstream?
promotes synthesis of macromolecules
46
growth in cells; what phase?
increased size of cell ⇒ G1 phase
47
proliferation
division
48
endoreplication cycle checkpoints (unconventional)
S and G results in polytenization
- no M and no cytokinesis
49
Polytenization
giant chromosomes with many stacks of replication on one another
- Ex: drosophila salivary gland, fat body, etc.
50
what is the degree of politely in drosophila?
1024-2048 DNA (2^10-11)
- 10-11 cycles of replication
51
glue proteins
multiple copies of genes allows a high level of gene expression ⇒ this can happen in a very short time frame
- Larva stage is a feeding time for drosophila but at the end they want to find a place to stick to a substrate with the glue protein
- Cells typically want to make a lot of proteins via transcriptional activation
52
Syncytium formation
occurs when there are replications of chromosomes and mitosis but no cytokinesis in 1 cell (Synchronous) ⇒ occurs in drosophila
Mitosis without cytokinesis
- S and M phase only (skips G)
53
what does syncytium result in?
thousands of nuclei in 1 cell ⇒ first 13 rounds mitosis (extremely rapid in 10 min intervals)
54
what cells in the human body become syncytial and how/
muscle cells become syncytial by fusing after being multiple cells
55
what kind of experiment would help you figure out how cells divide without killing the cell?
label chromosomal histones (H2A) with RFP as well as microtubule tubulin protein (spindles) with GFP
56
2 properties of cancer
1. Reproduce in defiance of the normal restraints on growth and division
2. Invade and colonize territories normally reserved for other cells
57
key properties of cancer (6)
1. Disregard signals that regulate cell proliferation
2. Avoid apoptosis
3. Escape replicative senescence and avoid differentiation
4. Genetically unstable
5. Invade surrounding tissues
6. Survive and proliferate in foreign sites (metastasis)
58
senescence
a phase of time when cells proliferate a lot before they stop and start differentiation
59
carcinomas
arise from epithelial cells
60
adenocarcinomas
arise from glandular tissue
61
sarcomas
arise from connective tissue or muscle cells
62
leukemias
from white blood cells and their precursors
63
lymphomas
derived from lymphocytes and found mainly within lymphoid organs
64
what are the majority of cancers?
epithelial cell cancers which lead to carcinomas
- There are also other types such as myelomas, leukemias, and lymphomas
65
neoplasm
tumor arising from the growth and proliferation of a cell in defiance of normal controls
66
benign
tumor is self limiting in growth and noninvasive ⇒ difference in invasion ability
- Usually these are easily removable
67
malignant
tumors have the ability to break away and travel via blood and lymph to other sites to form a metastasis
68
carcinogens
cause of mutations ⇒ chemicals, ionizing radiations (X-ray), viruses
- Types consist of chemical, physical, and biological
69
what initiates cancer?
mutations in the genes
70
most cancers can be traced to what?
a single abnormal cell
- A single mutation is not enough to cause a cancer
71
Chronic myelogenous leukemia (CML)
leukemia from white blood cells have the philadelphia chromosome created by a translocation between chromosome 9 and 22
- The site of breakage in all the leukemic cells is identical in any given patient indicating the cancer arises from a unique accident in a single cell
72
spontaneous mutation rate
10-6 mutations/gene*cell division => The number of divisions that occur over a lifespan is 1016
- Development of a cancer typically requires a substantial number of independent genetic mutations to occur in the lineage of one cell
73
T/F cancers increase with age
(T) The incidence of cancer increase with age as mutations accumulate
- cancer as a disease is age dependent
- peaks around 85 and then starts to decline again
74
when did global smoking peak? What was the lag time?
in 1990 and global lung cancer deaths caused by smoking followed after decades
- Phenomenon of tumor progression during this lag time where prospective cancer cells accumulated genetic and epigenetic changes
- The time needed to accumulate mutations explains the lag
75
what color are proliferating cells on a purple stain?
Proliferating cells are the dark stains
76
where are dividing cells in normal epithelium?
only in the basal layer
77
what are the progressive stages of an epithelial cancer?
1. normal epithelium
2. low grade intraepithelial neoplasia
3. high grade intraepithelial neoplasia
4. invasive carcinoma
- at first it is limited to the basal side of the epithelial tissue ⇒ eventually all epithelial layers are filled with actively dividing cells which invade and break the lamina into the connective tissue
78
what happens when cells have more mutations?
they predominantly grow due to natural selection
- Abnormal cells can become more and more abnormal and they grow more predominantly
79
heterogeneity
over time a variety of competing subclones arise in a tumor
- when they divide faster they have a further chance of speeding up division allowing them to become the dominant cell type => This also allows for them to become more mutated (mutagenesis selection)
80
what is cancer formation considered?
microevolution
81
genetic instability
mutations interfere with replication and genome maintenance leading to increases of mutation rates
- Defects in DNA repair ⇒ changes in DNA sequence, translocations and duplications, defects in chromosome segregation during mitosis ⇒ changes in karyotype
82
what happens when you hit a fundamental mechanism of genetic maintenance?
mutation becomes significant enough so you can’t maintain normal karyotypes
83
what cells are responsible for tissue maintenance in normal organ homeostasis?
stem cells
- Stem cells divide relatively slowly and undergoes asymmetric division to self renew and differentiate into multiple cell types (transit amplifying cells) ⇒ become different cells
- This creates heterogeneous cell populations
84
what cells do some cancers contain?
cancer stem cells
85
Cancer stem cells (CSCs)
maintain the tumor and large number of dividing cancer transit amplifying cells ⇒ derived from CSCs but have a limited capacity for self renewal
86
what happens when CSCs are purified and injected into an imunnodeficient animal?
they have a greatly enhanced ability to create new tumors in vivo
87
how do CSCs contribute to cancer returning?
many current cancer therapies preferentially kill the most rapidly proliferating cells, slow dividing cancer stem cells are less sensitive to these treatments
88
what kinds of mutations do cancer cells incur?
mutations that disable apoptotic mechanisms
89
what 2 things increase tumor growth?
1. an increase in cell division
2. a decrease in apoptosis
90
what do normal cells need to be attached to to survive? What about cancer cells?
normal cells do not divide unless they are attached to the substratum but cancer cells often divide in suspension
91
contact inhibition
normal cells stop moving and dividing when culture reaches confluence
92
transformed cancer phenotype
cancer cells continue dividing and pile up in layer upon layer in culture ⇒ when these cells change from normal they are transformed cells
93
foci
piles of uninhibited transformed (cancer) cells
94
where are cells extruded from in normal conditions?
cells are extruded into the lumen to generally undergo apoptosis
95
what happens when tumor cells are extruded apically
they might survive but are likely to be eliminated
96
what happens when tumor cells are extruded basally?
can more readily initiate invasion ⇒ this is different from extrusion on the apical side
97
what do normal adult cells do with glucose?
generally fully oxidize almost all glucose to produce ATP via oxidative phosphorylation
- Only when O2 is deprived, the normal cells convert pyruvate to lactate
98
what do cancer cells do with glucose?
they have abnormally increased glucose uptake and reduced oxidative phosphorylation (100-1000x more glucose uptake)
- This occurs regardless of oxygen presence
- This may be related to the condition of cancer where cells have a difficult time accessing oxygen via vessels
99
androgenesis
new blood cell formation created by cancers
100
Warburg effect (by-products?)
abnormally high glucose uptake where tumor cells form lactate and small molecule building blocks required for cell growth
- Byproducts like lactate which is building blocks may be helpful for cancer cells which facilitates rapid proliferation
101
what do tumor cells and supporting connective tissue/stroma rely on?
communication
- Cancer cells want oxygen and to clear waste through blood vessels
- It's not just vessels, but they also want to change their surrounding environments like the ECM
102
what do cancer cells create around them?
a microenvironment
103
tumor stroma
includes cancer cells, pericytes (vascular smooth muscle cells), fibroblasts, inflammatory white blood cells, blood and lymphatic vessels
- The tumor and its stroma evolve together
104
metastasis steps
1. break through the basal lamina
2. invade capillary
3. travel through bloodstream
4. adhere to the vessel wall
5. exit from the blood vessel
6. form metastasis at a different site
105
Epithelial mesenchymal transition (EMT)
cells are now floating (migrating) in connective tissues after breaking through the basal lamina to make them more mobile ⇒ epithelial like genes are no longer expressed and become more like mesenchymal genes
106
what are the 2 components of Epithelial mesenchymal transition
1. loosen cell-cell/cell-ECM adhesion ⇒ reduce both cadherins and integrins
2. Degrade ECM ⇒ secrete proteases and heparanases breaking into ECM
107
when can EMT happen aside from cancer?
EMT happens during normal development and regeneration ⇒ such as in drosophila
108
Circulating tumor cells (CTCs)
when tumor cells are traveling through the bloodstream after breaking the basal lamina
109
what happens when tumor cells exit from the blood vessel in their new location? what are the odds?
start proliferation at the new side inside of the new organ area
- This step is very difficult because cells know who they usually interact with and the new cells are very foreign
- The chance of the cells surviving at the foreign site is not that high
110
